The specificity of cellular immune responses in guinea pigs. I. T cells specific for 2,4 dinitrophenyl O tyrosyl residues

Guinea pigs immunized with the hapten 2,4 dinitrophenyl (DNP) coupled directly to Mycobacterium tuberculosis of strain H37Ra (DNP H37) show a variety of cell mediated immune responses to DNP coupled to protein carriers. The cells responsible for this specific response are thought to be T lymphocytes for the following reasons: guinea pigs immunized with DNP H37 displayed delayed hypersensitivity reactions to several DNP proteins and contact sensitivity to dinitrofluorobenzene; peritoneal exudate lymphocytes (PELs) obtained from DNP H37 immune animals respond to DNP proteins with DNP synthesis and cause inhibition of macrophage migration; PELs are highly enriched in T lymphocytes and contain few immunoglobulin bearing cells; further depletion of immunoglobulin bearing cells from this population does not diminish the in vitro proliferative response to antigen. Nitrophenyl conjugates of proteins lacking a paranitro group stimulated DNA synthesis poorly or not at all. In this respect, the specificity of T cells resembles that of DNP specific anitbody from the same animals. On the other hand, DNP conjugates of copolymers of glutamic acid and lysine and DNP conjugated to proteins via an interposed β alanyl glycyl glycyl spacer failed to stimulate DNA synthesis, although such compounds bind very efficiently to anti DNP antibody. By contrast, DNP conjugates of synthetic polypeptide carriers containing as little as 7% tyrosine strongly stimulated DNA synthesis in DNP H37 immune PELs. That the determinant responsible for this stimulation was DNP coupled to the hydroxyl group of tyrosine was shown by selective removal of DNP from tyrosine by thiolysis with 2 mercaptoethanol, which abolished their ability to stimulate T cells. This stimulatory capacity could be restored by redinitrophenylation, demonstrating that thiolysis had not simply inactivated the carrier. These data suggest that a major antigenic determinant recognized by such T cells is DNP coupled to the hydroxyl group of tyrosine. Nevertheless, either multivalent presentation or additional carrier structures are critical as mono O DNP L tyrosine and di O,N DNP L tyrosine failed to stimulate a proliferative response.